Abstract Toxicity and negative health effects of tobacco smoking results from the inhalation of a complex mixture of over 7000 chemicals including many toxicants and carcinogens included on the FDA list of Harmful and Potentially Harmful Constituents (HPHCs). Oxidants are a major class of toxicant abundant in tobacco smoke and are thought to play a critical role in the development of tobacco related diseases including COPD, CVD and cancer through the generation of oxidative stress/damage and inflammation. However, the specific oxidants most responsible remain unclear. While several oxidants are included on the HPHC list (eg. carbonyls), one of the most reactive, damaging and abundant classes, free radicals, are not represented. Recently we found that delivery of radicals varied greatly (>12-fold) across cigarette brands, was substantially impacted by product design (e.g. tobacco variety) and smoking behaviors and was high in other popular combustible products including little filtered cigars. Radical delivery was also highly correlated with other oxidants including carbonyls (eg. acrolein). Based upon these findings, we propose that tobacco-derived oxidants, including free radicals, represent an important target for developing regulatory strategies aimed at reducing the harm from combustible tobacco use. This approach is strongly supported by our preliminary findings that levels of oxidative stress biomarkers are significantly reduced in smokers who switch from high to low oxidant cigarettes. The objectives of this project are to identify specific oxidants responsible for the generation of tobacco related harm and determine the impact of oxidant reduction on tobacco-related toxicity endpoints. To this end, we propose 3 specific aims. In Aim 1, we will determine the levels and identity of free radicals and other oxidants delivered by different combustible tobacco products/brands using advanced electron paramagnetic resonance (EPR) spectroscopy and liquid chromatography-tandem mass spectrometry (LC- MS/MS) methodologies. In Aim 2 we conduct exposure studies in a relevant mouse model to determine the impact of tobacco smoke oxidants on lung damage and inflammation; comparing effects of high vs. low oxidant brands and tobacco varieties. In Aim 3, we will conduct mouse exposure studies to determine the impact of charcoal filtration of cigarette smoke on oxidant-induced lung damage in the mouse. Overall, these studies will significantly contribute to the field of tobacco regulatory science by focusing on the toxicological importance of oxidant exposure. These data will be of particular value to the FDA for the development of regulatory policies aimed at reducing harm imposed by tobacco usage.